Amino-terminal domain of NF1 binds to DNA as a dimer and activates adenovirus DNA replication.
NF1 is a DNA-binding protein involved in initiation of adenovirus DNA replication as well as in modulating the rate of transcription initiation of genes containing the sequence TGGCA. We show here that recombinant NF1 expressed via vaccinia virus is transported into the nucleus and binds to its cognate sequences with the same specificity as NF1 purified from HeLa cells. Furthermore, the recombinant NF1 forms oligomers in solution and binds as a dimer to palindromic as well as half-site sequences. NF1 expressed via vaccinia virus stimulates the initiation of adenovirus replication in vitro. The N-terminal 240 amino acids of the protein are sufficient for full DNA-binding activity as well as stimulation of adenovirus replication. By analysis of several NF1 mutants translated in vitro, we also define the minimal DNA-binding domain and localize the region responsible for DNA binding on the N-terminal and for oligomerization on the C-terminal side of this domain.
Gounari
F
F
European Molecular Biology Laboratory, Heidelberg, FRG.
De Francesco
R
R
Schmitt
J
J
van der Vliet
P
P
Cortese
R
R
Stunnenberg
H
H
D000260:Adenoviruses, Human / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001667:Binding, Competitive; D022762:CCAAT-Enhancer-Binding Proteins; D004261:DNA Replication; D004279:DNA, Viral / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D015345:Oligonucleotide Probes; D010957:Plasmids; D014176:Protein Biosynthesis; D011994:Recombinant Proteins / Q000378:metabolism; D011995:Recombination, Genetic; D014157:Transcription Factors; D014158:Transcription, Genetic; D014616:Vaccinia virus / Q000235:genetics; D051840:Y-Box-Binding Protein 1
D022762:CCAAT-Enhancer-Binding Proteins; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D015345:Oligonucleotide Probes; D011994:Recombinant Proteins; D014157:Transcription Factors; D051840:Y-Box-Binding Protein 1; C070856:YBX1 protein, human